By using optical tweezers with an adjustable trap stiffness, we have performed systematic single molecule stretching experiments with two types of DNA-intercalator complexes, in order to investigate the effects of the maximum applied forces on the mechanical response of such complexes. We have explicitly shown that even in the low-force entropic regime the persistence length of the DNA-intercalator complexes is strongly force-dependent, although such behavior is not exhibited by bare DNA molecules. We discuss the possible physicochemical effects that can lead to such results. In particular, we propose that the stretching force can promote partial denaturation on the highly distorted double-helix of the DNA-intercalator complexes, which interfere strongly in the measured values of the persistence length.

We have performed a biophysical characterization, at single molecule level, of the interaction between the DNA molecule and the biogenic polyamine putrescine. By using force spectroscopy, we were able to monitor the complexes formation as putrescine is added to the sample, determining the mechanical properties of such complexes and the physicochemical (binding) parameters of the interaction for three different ionic strengths. In particular, it was shown that the behavior of the equilibrium binding constant as a function of the counterion concentration deviates from the prediction of the Record-Lohman model. The measured constants were (1.3 ± 0.2) × 10 M for [Na] = 150 mM, (2.1 ± 0.2) × 10 M for [Na] = 10 mM, and (2.2 ± 0.3) × 10 M for [Na] = 1 mM. The cooperativity degree of the binding reaction, on the other hand, increases with the ionic strength. From these analysis, the DNA-putrescine binding mechanisms are inferred, and a comparison with results reported for ordinary bivalent ions like magnesium is performed. Such study provides new insights on the general behavior of the DNA interactions with biogenic polyamines.

In this work, we have measured, by means of optical tweezers, forces acting on depletion-induced DNA condensates due to the presence of the DNA-like charged protein bovine serum albumin (BSA). The stretching and unfolding measurements performed on the semi-flexible DNA chain reveal (1) the softening of the uncondensed DNA contour length and (2) a mechanical behavior strikingly different from those previously observed: the force-extension curves of BSA-induced DNA condensates lack the "saw-tooth" pattern and applied external forces as high as ≈80 pN are unable to fully unfold the condensed DNA contour length. This last mechanical experimental finding is in agreement with force-induced "unpacking" detailed Langevin dynamics simulations recently performed by Cortini et al. on model rod-like shaped condensates. Furthermore, a simple thermodynamics analysis of the unfolding process has enabled us to estimate the free energy involved in the DNA condensation: the estimated depletion-induced interactions vary linearly with both the condensed DNA contour length and the BSA concentration, in agreement with the analytical and numerical analysis performed on model DNA condensates. We hope that future additional experiments can decide whether the rod-like morphology is the actual one we are dealing with (e.g. pulling experiments coupled with super-resolution fluorescence microscopy).

No abstract